The long term goal of this research is to develop a microfluidic platform to investigate the intercommunication of nerve cells in networks and the influence of the interconnections between cells on the neuronal response to physical trauma and exposure to chemical toxins. A key aspect of this work will be to develop and utilize three-dimensional microfluidic systems to precisely control delivery of reagents to individual cells in a network and to use this system to further our understanding of cell-to-cell communication. Cell networks in vitro will be used to model complex communication in the larger network of the brain. Special emphasis will be placed on understanding pre- vs. postsynaptic receptors, plasticity in patterns and screening for pharmacological efficacy with multiple cells at once. It is normally difficult to carry out detailed studies of drugs, toxins or damage to specific cells in neural networks owing to an inability to simultaneously stimulate (or expose to drug, toxin) and record from several specific cells in an array or network. The system proposed will provide a solution to this problem. We expect the work will expand existing knowledge concerning molecule-based cell-to-cell communication pathways and provide new avenues for screening drugs and toxins involved in the brain. There are four specific aims for this proposal. First, an integrated system will be designed and fabricated to allow linear cell arrays to be non-invasively stimulated and continuously monitored in real-time. Second, a three-dimensional microfluidic device combining small apertures with cross flow will be used to address more complicated two-dimensional cell networks. Third, the microfluidic system combined with fluorescence monitoring of cell activity will be tested on PC12 and P19 cell networks as models. Fourth, neuronal cell cultures with functional synapses will be monitored. The long-term goal is to use this system to examine the effects of physical and chemical damage to specific cells in a neuronal network. We propose to test the hypotheses that 1) neuronal connectivity changes to compensate for neuronal loss following physical damage, 2) activity in neuronal networks affects the rate of degradation following exposure to toxins, 3) glutamate neurons play an active role in dopaminergic neuronal cell loss following exposure toxins, and 4) L-DOPA plays a role in the progression of cell loss in Parkinson's Disease and might be involved in the mechanism of action of neuroprotective agents on dopamine cells. The proposed research has the overarching goal of establishing microfluidics as a valuable tool for biology.